Insulin stimulates glucose transport in rat adipose cells through the translocation of glucose transporters from an intracellular pool to the plasma membrane. A detailed characterization of the subcellular fractions of these cells, and of the distribution of glucose transporters among these fractions is now described. The results suggest that the intracellular pool of glucose transporters may comprise a specialized membrane species and that intracellular glucose transporters may be processed during their cycling to the plasma membrane in response to insulin. The effects of TRIS on insulin's stimulation of glucose transport have been investigated. The results suggest that insulin's action on the translocation of glucose transporters may occur at a step whereby glucose transporters associated with the plasma membrane become functional. The effects of low temperature on the stimulation of glucose transport by insulin have been examined. The results confirm that insulin appears to stimulate glucose transport through a step in which glucose transporters in the plasma membrane become functional. The mechanism of insulin resistant glucose transport in isolated guinea pig adipocytes has been examined. The results suggest that this resistance is the consequence of a relative depletion in the number of intracellular glucose transporters. In a preliminary series of experiments, insulin appears to stimulate glucose transport in isolated human adipose cells by a translocation mechanism similar to that observed in rat adipose cells and diaphragm. The photoaffinity crosslinking agent hydroxysuccinimidyl-4-azidobenzoate has been used to attach [3H]cytochalasin b to a rat adipocyte low-density microsomal membrane protein of 45-50 kDa. The characteristics of this protein are consistent with those of the glucose transporter. Thus, the identification, isolation, and purification of the adipocyte glucose transporter should now be possible.